Analysis of proteins in histological sections and other cytological preparations may be performed using fluorescence in situ hybridization (FISH). FISH is a cytogenetic technique for detecting and localizing the presence or absence of specific DNA sequences on chromosomes. FISH uses fluorescent probes that bind to only those parts of the chromosome with which they show a high degree of sequence complementarity. Fluorescence microscopy can be used to find out where the fluorescent probe is bound to the chromosomes by counting the fluorescent dots present in an image.
For FISH analysis in carcinoma tumor samples, the measurement of the FISH dot counts is relevant only for the epithelial cells in a sample. However, conventional FISH analysis measures the FISH signal for all cells in a field of view because there lacks an effective way of limiting the analysis to a defined subset of cells. In the absence of manual intervention, this can lead to many non-epithelial cells (such as stromal cells) contributing to the FISH dot counting statistics, which leads to less accurate dot counting measurements.